R-type bacteriocins are minimal contractile nanomachines that hold promise as precision antibiotics 1 – 4 . Each bactericidal complex uses a collar to bridge a hollow tube with a contractile sheath loaded in a metastable state by a baseplate scaffold 1 , 2 . Fine-tuning of such nucleic acid-free protein machines for precision medicine calls for an atomic description of the entire complex and contraction mechanism, which is not available from baseplate structures of (DNA-containing) T4 bacteriophage 5 . Here we report the atomic model of the complete R2 pyocin in its pre- and post-contraction states, each containing 384 subunits of 11 unique atomic models of 10 gene products. Comparison of these structures suggests the sequence of events during pyocin contraction: tail fibers trigger lateral dissociation of baseplate triplexes; the dissociation then initiates a cascade of events leading to sheath contraction; this contraction converts chemical energy into mechanical force to drive the iron-tipped tube across the bacterial cell surface, killing the bacterium.

Listeria monocytogenes is a significant foodborne human pathogen that can cause severe disease in certain high-risk individuals. L. monocytogenes is known to produce high-molecular-weight, phage tail-like bacteriocins, or "monocins," upon induction of the SOS system. In this work, we purified and characterized monocins and found them to be a new class of F-type bacteriocins. The L. monocytogenes monocin genetic locus was cloned and expressed in Bacillus subtilis, producing specifically targeted bactericidal particles. The receptor binding protein, which determines target cell specificity, was identified and engineered to change the bactericidal spectrum. Unlike the F-type pyocins of Pseudomonas aeruginosa, which are related to lambda-like phage tails, monocins are more closely related to TP901-1-like phage tails, structures not previously known to function as bacteriocins. Monocins therefore represent a new class of phage tail-like bacteriocins. It appears that multiple classes of phage tails and their related bacteriocins have coevolved separately in parallel. IMPORTANCEPhage tail-like bacteriocins (PTLBs) are structures widespread among the members of the bacterial kingdom that are evolutionarily related to the DNA delivery organelles of phages (tails). We identified and characterized "monocins" of Listeria monocytogenes and showed that they are related to the tail structures of TP901-1-like phages, structures not previously known to function as bacteriocins. Our results show that multiple types of envelope-penetrating machines have coevolved in parallel to function either for DNA delivery (phages) or as membrane-disrupting bacteriocins. While it has commonly been assumed that these structures were coopted from phages, we cannot rule out the opposite possibility, that ancient phages coopted complex bacteriocins from the cell, which then underwent adaptations to become efficient at translocating DNA.T wo types of high-molecular-mass (Ͼ10 6 -Da), phage tail-like bacteriocins (PTLBs), the R type and the F type, are known to exist in the bacterial kingdom (1, 2). R-type bacteriocins (RTBs) are contractile nanotube machines evolutionarily related to Myoviridae tails (e.g., that of bacteriophage T4) (3), type 6 secretion systems (4, 5), insecticidal protein injector complexes (6, 7), and structures involved in bacterium-marine animal interactions (8). RTBs are composed of a tube surrounded by a contractile sheath (9). At one end of the sheath is a complex baseplate structure to which six receptor binding proteins (RBPs) are attached. RTBs kill target bacteria by first binding to a receptor on the cell surface via RBPs. This event triggers sheath contraction, which then drives the tube structure through the cell envelope. The result is a dissipation of the membrane potential and cell death. Contact with a single RTB particle is sufficient to kill a cell (10). Far less studied are the F-type bacteriocins (FTBs). These high-molecular-weight bactericidal protein structures are evolutionarily related to the nonco...